Modulation of Cardiac Function by Myosin Light Chain Kinases and Phosphatases

肌球蛋白轻链激酶和磷酸酶对心脏功能的调节

基本信息

批准号：
9919619
负责人：
Audrey N Chang
金额：
$ 40.5万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-05-01 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9919619
关键词：
Ablation Acute Adult Affect Animal Model Biochemical Biological Assay Cardiac Cardiac Myocytes Cardiac Myosins Cardiomyopathies Catalytic Domain Cause of Death Chemosensitization DNA Sequence Alteration Dilated Cardiomyopathy Dobutamine Enzymes Family member Functional disorder Future Genes Genetic Goals Heart Heart Abnormalities Heart Diseases Heart Transplantation Heart failure Human Hypertrophy Infusion procedures Investigation Knock-out Knockout Mice Light MYLK gene Maintenance Measures Mediating Modeling Molecular Motors Mus Muscle Muscle function Myocardium Myopathy Myosin ATPase Myosin Light Chain Kinase Myosin Regulatory Light Chains Patients Performance Pharmacology Pharmacotherapy Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Site Phosphotransferases Physiological Prevalence Property Propranolol Protein Dephosphorylation Protein Kinase Proteins Resistance Role Sarcomeres Signal Pathway Signal Transduction Site System Testing Therapeutic Time Transcript cardiac muscle disease conditional knockout heart function improved in vivo inorganic phosphate insight myosin phosphatase overexpression

Ablation Acute Adult Affect Animal Model Biochemical Biological Assay Cardiac Cardiac Myocytes Cardiac Myosins Cardiomyopathies Catalytic Domain Cause of Death Chemosensitization DNA Sequence Alteration Dilated Cardiomyopathy Dobutamine Enzymes Family member Functional disorder Future Genes Genetic Goals Heart Heart Abnormalities Heart Diseases Heart Transplantation Heart failure Human Hypertrophy Infusion procedures Investigation Knock-out Knockout Mice Light MYLK gene Maintenance Measures Mediating Modeling Molecular Motors Mus Muscle Muscle function Myocardium Myopathy Myosin ATPase Myosin Light Chain Kinase Myosin Regulatory Light Chains Patients Performance Pharmacology Pharmacotherapy Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Site Phosphotransferases Physiological Prevalence Property Propranolol Protein Dephosphorylation Protein Kinase Proteins Resistance Role Sarcomeres Signal Pathway Signal Transduction Site System Testing Therapeutic Time Transcript cardiac muscle disease conditional knockout heart function improved in vivo inorganic phosphate insight myosin phosphatase overexpression

展开

项目摘要

PROJECT SUMMARY Cardiac contraction depends on the molecular motor myosin in sarcomeres where maintenance of contractile performance is achieved in part by the constitutive phosphorylation of myosin regulatory light chain (RLC) through the respective activities of myosin light chain kinase (MLCK) and phosphatase (MLCP). Dilated cardiac myopathy in mice and humans results in decreased cardiac MLCK (cMLCK) expression and RLC phosphorylation while animal models with increased phosphorylation have enhanced cardiac performance with resistance to heart failure. Although it is well established that RLC phosphorylation is important for normal cardiac function, surprisingly little is known about signaling mechanisms regulating cMLCK and MLCP activities, respectively. We propose investigations on cardiac-specific signaling mechanisms for these two enzymes that affect myosin phosphorylation to elucidate cellular mechanisms for normal function as well as potential causes of decreased RLC phosphorylation in heart failure. Specific Aim 1: Having recently discovered cMLCK is phosphorylated in vivo, we plan to identify roles of specific phosphorylation sites in regulating cMLCK activity using cMLCK expression and assay systems I developed to test the hypothesis that cMLCK phosphorylation enhances its activity. These studies will be extended to intact cardiac muscle to define signaling mechanisms involved in cMLCK phosphorylation, including responsible protein kinases. Additionally, we will test the hypothesis that other forms of heart failure involve reduced RLC phosphorylation to identify potentially common signaling derangements. Specific Aim 2. Determine the roles of myosin-targeted and soluble phosphatase activities in mediating RLC dephosphorylation. Using conditional knockout models for cardiac MYPT2 and the related, ubiquitously expressed subunit MYPT1 in adult mice, we will assess the effects of specific gene ablation on cardiac function. Intact cardiac muscle trabeculae from wildtype and knockout hearts will be used to quantitatively measure contributions of distinct pools of phosphatases to maintenance of half-maximal RLC phosphorylation. Cardiomyocytes from MYPT1 and MYPT2 as well as PP1cδ knockout mice will be used to identify the regulatory subunit for the soluble phosphatase. These studies will test the hypothesis that PP1cδ bound and unbound to MYPT2 specifically dephosphorylates RLC, providing insights into the physiological role of myosin phosphatases in the heart. These results will also set the stage for future studies on aberrant signaling pathways that cause cardiac muscle dysfunction through effects on RLC phosphorylation.

项目摘要心脏收缩取决于肉瘤中的分子运动肌球蛋白，在那里保持收缩。肌球蛋白调节轻链（RLC）的组成型磷酸化可以部分实现性能通过肌球蛋白轻链激酶（MLCK）和磷酸酶（MLCP）的相关活性。心脏扩张小鼠和人类的肌病导致心脏MLCK（CMLCK）和RLC改善磷酸化虽然磷酸化增加的动物模型具有增强的心脏性能抵抗心力衰竭。尽管已经很好地确定RLC磷酸化对于正常心脏很重要功能，关于调节CMLCK和MLCP活动的信号传导机制知之甚少，分别。我们提出了有关这两种酶的心脏特异性信号传导机制的研究影响肌球蛋白磷酸化以阐明正常功能以及潜在原因的细胞机制心力衰竭中RLC磷酸化的降低。特定目标1：最近发现CMLCK是磷酸化的体内，我们计划确定特定磷酸化位点在调节CMLCK活性中的作用我开发了使用CMLCK表达和测定系统来检验CMLCK磷酸化的假设增强其活性。这些研究将扩展到完整的心肌以定义信号传导机制参与CMLCK磷酸化，包括负责的蛋白激酶。此外，我们将测试假设其他形式的心力衰竭涉及降低RLC磷酸化以鉴定潜在的常见信号演变。具体目标2。确定靶向肌球蛋白的作用和可溶性磷酸酶的作用介导RLC去磷酸化的活性。使用有条件的淘汰模型用于心脏MyPT2和相关的，普遍表达的成年小鼠中的亚基MYPT1，我们将评估特定基因消融的影响关于心脏功能。野生型和淘汰心脏的完整心肌小梁将用于定量测量不同磷酸酶池对维持半最大RLC的贡献磷酸化。来自MyPT1和MyPT2以及PP1CΔ基因敲除小鼠的心肌细胞将用于确定固体光肽酶的调节亚基。这些研究将检验PP1Cδ的假设绑定并未与MYPT2绑定并专门脱磷酸化RLC，提供了对身体角色的见解心脏中肌球蛋白磷酸酶这些结果还将为以后的关于异常信号传导的研究奠定阶段通过对RLC磷酸化的影响引起心肌功能障碍的途径。